大黄灵仙胶囊含药血清对胆管细胞炎症模型miRNA⁃140⁃5p表达及NF-κB/NLRP3信号通路的影响  

Effect of drug-containing serum of Dahuang Lingxian Capsule on miRNA-140-5p expres⁃sion and NF-κB/NLRP3 signaling pathway in bile duct cell inflammation model

作　　者：居燕飞 陈雅璐 林龙 劳永彩 王清坚[2] JU Yan-fei;CHEN Ya-lu;LIN Long;LAO Yong-cai;WANG Qing-jian(Guangxi University of Chinese Medicine,Nanning 530200,China;First Affiliated Hospital of Guangxi University of Chinese Medicine,Nanning 530200,China)

机构地区：[1]广西中医药大学,广西南宁530200 [2]广西中医药大学第一附属医院,广西南宁530023

出　　处：《时珍国医国药》2025年第4期628-635,共8页Lishizhen Medicine and Materia Medica Research

基　　金：国家自然科学基金(82060867,82474507);广西自然科学基金(2024GXNSFDA010025,2024GXNSFBA010106);广西重点研发计划项目(桂科AB24010130);广西名中医传承工作室建设项目(GZY2024011);广西壮族自治区研究生教育创新计划项目(YCSW2022350,YCSY2023037)。

摘　　要：目的探讨大黄灵仙胶囊(DHLX)含药血清对脂多糖(LPS)诱导的大鼠胆管细胞炎症模型miRNA-140-5p表达及NF-κB/NLRP3信号通路的可能作用机制。方法12只SD大鼠随机分为空白血清组和含药血清组,每组6只。含药血清组给予320 mg·kg^(-1)·d^(-1)灌胃,空白血清组给予等量蒸馏水灌胃,连续3 d,每天1次,末次给药6 h后腹主动脉取血制备含药血清。将胆管细胞分为空白组、模型组、DHLX组、DHLX+抑制剂组、DHLX+抑制剂对照组。收集上述各组细胞,电镜观察胆管细胞形态改变;检测各组细胞半胱氨酸蛋白酶-1(Caspase-1)活性;ELISA测定白细胞介素1β(IL-1β)、白细胞介素18(IL-18)浓度;qRT-PCR检测各组细胞miRNA-140-5p以及肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)、Toll样受体4(TLR4)、核因子κB p65(NF-κB p65)、NOD样受体热蛋白结构域相关蛋白3(NLRP3)、Caspase-1、凋亡相关斑点样蛋白(ASC)mRNA表达量;Western blot检测TNF-α、IL-6、TLR4、NF-κB p65、NLRP3、Caspase-1、ASC蛋白表达水平。结果与空白组比较,模型组胆管细胞核固缩、胞膜出泡、胞质空泡化,线粒体嵴模糊不清、减少或消失,内质网膨胀,Caspase-1活性表达显著增加(P<0.01),IL-1β、IL-18,TNF-α、IL-6、TLR4、NF-κB p65、NLRP3、Caspase-1、ASC mRNA及蛋白表达增加(P<0.05),miRNA-140-5p表达显著降低(P<0.01);与模型组比较,DHLX组胆管细胞损伤减轻,Caspase-1活性表达显著降低(P<0.01),IL-1β、IL-18,TNF-α、IL-6、TLR4、NF-κB p65、NLRP3、Caspase-1、ASC mRNA及蛋白表达降低(P<0.05),miRNA-140-5p表达显著增加(P<0.01);与DHLX组比较,DHLX+抑制剂组胆管细胞损伤加重,Caspase-1活性表达显著增加(P<0.01),IL-1β、IL-18,TNF-α、IL-6、TLR4、NF-κB p65、NLRP3、Caspase-1、ASC mRNA及蛋白表达增加(P<0.05),miRNA-140-5p表达显著降低(P<0.01),DHLX+抑制剂对照组上述指标差异均无统计学意义(P>0.05);与DHLX+抑制剂组比较,DHLX+抑制剂对照组胆管细胞焦亡情Objective To explore the potential mechanisms of drug-containing serum of Dahuang Lingxian Capsule(DHLX)on miR⁃NA-140-5p expression and the NF-κB/NLRP3 signaling pathway in bile duct cells induced by lipopolysaccharide(LPS)in a rat model of inflammationMethods Twelve SD rats were randomly divided into the blank serum group and drug-containing serum group,with 6 rats in each group.The drug-containing serum group was given 320 mg·kg^(-1)·d^(-1) intragastric administration,while the blank serum group received an equal volume of distilled water.Treatment was given once daily for 3 consecutive days,and the drug-containing serum was prepared by taking blood from abdominal aorta 6 hours after the final administration.Bile duct cells were divided into the blank group,model group,DHLX group,DHLX+inhibitor group and DHLX+inhibitor control group.Cells from each group were col⁃lected,and the morphological changes in bile duct cells were observed under electron microscope.Cell Caspase-1 activity was detected in each group.Levels of interleukin-1β(IL-1β)and interleukin-18(IL-18)were determined by ELISA.qRT-PCR was used to determine the expression levels of miRNA-140-5p,tumor necrosis factorα(TNF-α),interleukin-6(IL-6),Toll-like receptor 4(TLR4),nuclear factorκB p65(NF-κB p65),NOD-like receptor pyrin domain-containing protein 3(NLRP3),Caspase-1,and apoptosis-associated speck-like protein(ASC)mRNA.Western blot(WB)was performed to detect the protein expression levels of TNF-α,IL-6,TLR4,NF-κB p65,NLRP3,Caspase-1,and ASC.Results Compared with blank group,the model group exhib⁃ited nuclear contraction,cell membrane vesiculation,cytoplasmic vacuolation,blurred or reduced mitochondrial cristae,endoplasmic re⁃ticulum expansion,and Caspase-1 activity was significantly increased(P<0.01).The mRNA and protein expressions of IL-1β,IL-18,TNF-α,IL-6,TLR4,NF-κB p65,NLRP3,Caspase-1 and ASC were increased(P<0.05),while the expression of miR⁃NA-140-5p was significantly decreased(P<0.01).Compared with the model group,bile d

关 键 词：胆石症 胆管细胞 大黄灵仙胶囊 miRNA-140-5p NF-κB/NLRP3信号通路 

分 类 号：R285.5[医药卫生—中药学]